1. Field of the Invention
The present invention relates to improved methods for the treatment of metabolic syndrome, which includes diabetes, cardiovascular disease, hyperlipidemia, liver steatosis, and obesity. More particularly, provided herein are glycosylation isoforms of TSH receptor (TSHR) agonists that exhibit increased bioactivity on adipose tissue for treatment of metabolic syndrome, and yet avoid thyrotoxicosis, the chronic hyperthyroid state.
2. Description of the Related Art
Metabolic syndrome is a public health problem that is both serious and widespread. Metabolic syndrome in humans is typically associated with obesity and characterized by one or more of the following: cardiovascular disease, liver steatosis, hyperlipidemia, diabetes, and insulin resistance. One third of the population in industrialized countries has an excess weight of at least 20% relative to the ideal weight. This phenomenon has spread to the developing world, particularly to the regions of the globe where economies are modernizing. As of the year 2000, there were an estimated 300 million obese people worldwide. Obesity is frequently attributed to consumption of a high fat diet, inactivity, and genetic predisposition.
Obesity considerably increases the risk of developing cardiovascular or metabolic diseases. For an excess weight greater than 30%, the incidence of coronary disease is doubled in subjects less than 50 years of age. Studies carried out for other diseases are equally revealing. For an excess weight of 20%, the risk of high blood pressure is doubled. For an excess weight of 30%, the risk of developing non-insulin dependent diabetes is tripled, and the incidence of dyslipidemia increases six fold. The list of additional diseases promoted by obesity is long; abnormalities in hepatic function, digestive pathologies, certain cancers, and psychological disorders are prominent among them.
Treatments for obesity include restriction of caloric intake and increased caloric expenditure through physical exercise. However, the treatment of obesity by dieting, although effective in the short-term, suffers from an extremely high rate of recidivism. Treatment with exercise has been shown to be relatively ineffective when applied in the absence of dieting. Other treatments include gastrointestinal surgery or agents that limit the absorption of dietary lipids. These strategies have been largely unsuccessful due to side effects of their use. Current therapies for complications associated with obesity, including type-2 diabetes, hyperlipidemia, and steatohepatitis, have been inadequate to halt the progression of these life-threatening pathologies in most instances.
Lipolysis is the biochemical process by which stored fats in the form of triglycerides are released from fat cells as individual free fatty acids (FFA) into the circulation. Stimulation of lipolysis has been clearly linked to increased energy expenditure in humans, and several strategies to promote lipolysis and increase oxidation of lipids have been investigated to promote weight loss and treat aspects of metabolic syndrome associated with obesity. These therapeutic efforts primarily focus on creating compounds that stimulate the sympathetic nervous system (SNS) through its peripheral β-adrenoreceptors.
Lipolytic agents have been investigated extensively in rodents, dogs, and primates and found to produce striking improvements in adiposity, glucose sensitivity, and dyslipidemia, hallmarks of the metabolic syndrome. These agents, agonists of sympathetic nervous system catecholamines, have not proven to be successful therapeutics in humans principally due to the inability thus far to create specific agents that target only adipose tissue without stimulating other tissues responsive to sympathetic innervation.
Energy expenditure represents one side of the energy balance equation. In order to maintain stable weight, energy expenditure should be in equilibrium with energy intake. Considerable efforts have been made to manipulate energy intake (i.e., diet and appetite) as a means of maintaining or losing weight; however, despite enormous sums of money devoted to these approaches, they have been largely unsuccessful. There have also been efforts to increase energy expenditure pharmacologically as a means of managing weight control and treating obesity. Increasing metabolic rate is an attractive therapeutic approach because it has the potential of allowing affected individuals to maintain food intake at normal levels. Further evidence supports the view that increases in energy expenditure due to pharmacological means are not fully counteracted by corresponding increases in energy intake and appetite. See Bray, G. A. (1991) Ann Rev Med 42, 205-216.
Much of the energy expended on a daily basis derives from resting metabolic rate (RMR), which comprises 50-80% of the total daily energy expenditure. For a review, see Astrup, A. (2000) Endocrine 13, 207-212. Noradrenaline turnover studies have shown that most of the variability in RMR that is unexplained by body size and composition is related to differences in SNS activity, suggesting that SNS activity does modulate RMR. See Snitker, S., et al. (2001) Obes. Rev. 1:5-15. Meal ingestion is accompanied by increased SNS activity, and studies have demonstrated that increased SNS activity in response to a meal accounts for at least part of meal-induced thermogenesis.
The peripheral targets of the SNS involved in the regulation of energy utilization are the β-adrenoreceptors (β-AR's). These receptors are coupled to the second messenger cyclic adenosine monophosphate (cAMP). Elevation of cAMP levels leads to activation of protein kinase A (PKA), a multi-potent protein kinase and transcription factor eliciting diverse cellular effects. See Bourne, H. R., et al. (1991) Nature 349:117-127. Adipose tissue is highly innervated by the SNS, and possesses three known subtypes of β-adrenoreceptors, β1-, β2-, and β3-AR. Activation of the SNS stimulates energy expenditure via coupling of these receptors to lipolysis and fat oxidation. Increased serum free fatty acids (FFAs) produced by adipose tissue and released into the bloodstream stimulate energy expenditure and increase thermogenesis. For a review, see Astrup, A. (2000) Endocrine 13, 207-212. In addition, elevated PKA levels increase energy utilization in fat by up-regulating uncoupling protein-1 (UCP-1), which creates a futile cycle in mitochondria, generating waste heat.
Over the past two decades, investigation of the physiological benefits of SNS activation for the treatment of obesity and treatment of diabetes related to obesity has centered on pharmacological activation of the β3-AR. Expression of the β3-AR is restricted to a narrower range of tissues than the β1 or β2 isoforms, and is highly expressed in rodent adipose tissue compared to the other isoforms. Experimental work in rodents treated with β3-AR agonists has demonstrated that stimulation of lipolysis and fat oxidation produces increased energy expenditure, weight loss, and increased insulin sensitivity. See de Souza, C. J. and Burkey, B. F. (2001) Curr. Pharm. Des. 7, 1433-1449. However, the potential benefits of the β3-AR agonists have not been realized, due to their lack of efficacy at the human β3-AR. Further, it has more recently been shown that the levels of β3-AR in rodent adipose tissue are much higher than in human adipose tissue. In human adipose tissue, the β1 and β2 isoforms represent the predominant adrenoreceptor isoforms. See Arch, J. R. (2002) Eur. J. Pharmacol. 440:99-107. Thus, although stimulation of lipolysis has been demonstrated in rodents, the mechanism for therapeutically producing the corresponding effects in humans is unrealized.
Energy expenditure can be stimulated pharmacologically by manipulation of the central nervous system, by activation of the peripheral efferents of the SNS, or by increasing thyroid hormone levels. Thyroid hormone stimulates carbohydrate and lipid catabolism in most cells of the body and increases the rate of protein synthesis. Thyroid-stimulating hormone (TSH) stimulates thyroid hormone biosynthesis and secretion. The secretion of TSH from the thyrotrophs of the anterior pituitary is inhibited by circulating T4 and T3 and stimulated by thyrotropin-releasing hormone (TRH) produced in the hypothalamus. See Utiger, in. Endocrinology and Metabolism (Felig and Frohman, eds), pp. 261-347, McGraw-Hill, (2001). The hypothalamic-pituitary-thyroid (HPT) axis is a classical endocrine feedback pathway negatively regulated by thyroid hormone T3, which is released by the thyroid gland or synthesized in tissues from T4, the other form of thyroid hormone released by the thyroid gland. Release of hypothalamic TRH is inhibited by T3, and synthesis of pituitary TSH is inhibited by T3.
As a result of the catabolism produced by thyroid hormone, heat is given off and energy expenditure is increased. There has been an intense interest in thyroid hormone levels in obesity, due to the opportunity to increase basal energy consumption by increasing thyroid hormone levels. Studies of thyroid tissue have revealed that the thyroid receives persistent stimulation with TSH. The thyroid is a slow-reacting organ, with thyrocytes requiring sustained 18-hour TSH stimulation in order to initiate DNA synthesis and proliferation. See Roger, P. et al. (1987) J. Cell Physiol. 130, 58-67.
Recombinant human TSH (rhTSH) has been introduced into humans (Thyrogen®, Genzyme Corporation, Cambridge, Mass.) and has a much lower metabolic clearance rate (MCR) than human pituitary-derived TSH. Estimates of the mean apparent elimination half-life are 25+/−10 hours. Serum concentrations of rhTSH are significantly elevated up to 24 hours after a single injection of approximately 30 μg/kg in human subjects. See Ladenson, P. W. et al. (1997) N. Engl. J. Med. 337, 888-896. Human pituitary TSH is a glycoprotein mixture of oligosaccharide isoforms, including sulfated oligosaccharides, sialylated oligosaccharides, and oligosaccharides that lack anionic groups. In the hypothyroid state, sustained exposure to TSH is needed to increase thyroid hormone release and sialylated TSH produces greater in vivo thyroid-stimulating activity than other TSH glycoforms.
However, studies have revealed that obese and normal-weight individuals have similar thyroid hormone profiles. An excess of thyroid hormone leads to various disorders, generally termed thyrotoxicosis. This condition is characterized by an abnormally high metabolic rate, increased blood pressure, high body temperature, heat intolerance, irritability, and tremors of the fingers. Of particular concern in the obese state is the tendency toward increased and more forceful heartbeats. Due to the adverse effects of elevated thyroid hormone levels, the use of thyroid hormone to treat obesity has seen little success, other than in the small fraction of obese patients identified with hypothyroidism.
Clearly a need remains for improved treatments that are useful for stimulating lipolysis and treating metabolic syndrome without producing potentially serious side effects associated with the hyperthyroid state. The present invention fulfills such needs and offers other related advantages.